Phospholipids are essential for membrane barrier function, as precursors to other cellular components, and as important regulatory molecules and cofactors in enzyme catalyzed reactions. Due to this pleiotropic nature for the requirement of phospholipids, it has been difficult to precisely establish the relationships between membrane phospholipid composition and cell functions and the specific role of each phospholipid. A combined biochemical and molecular genetic approach will be carried out in Escherichia coli to investigate the role of phospholipids in cell function. The genes encoding the enzymes which are responsible for the synthesis of phosphatidylethanolamine, phosphatidylserine, phosphatidylglycerol and cardiolipin will be placed under the control of promoters such as those of the lactose and arabinose operons which will place the expression of the "phospholipid genes" under the exogenous regulation of the concentration in the growth medium of the inducers of these operons. This will allow the systematic regulation of the level of each of the above phospholipids. Gene products which are responsible for regulating the ratios of phospholipid to membrane protein and anionic to zwitterionic phospholipids will also be similarly regulated. The effect on membrane functions such as transport of specific solutes, cell motility, phage infectivity, colicin susceptibility, protein translocation across the inner membrane, membrane potential of whole cells and isolated membrane vesicles, cell morphology, and osmotic stability will be correlated with phospholipid content and composition. Conditions (genetic or metabolic) which suppress or reduce the requirement for various phospholipids will be sought. Specific functions for which there is some evidence of phospholipid involvement will be studied as a function of the level and composition of phospholipids. Anionic phospholipids have been implicated in protein translocation across the cytoplasmic membrane, the insertion of colicins into membranes, and the initiation of DNA synthhesis. Phosphatidylethanolamine has been implicated in the assembly of the lactose and proline permeases, membrane permeability properties, and the synthesis of the core region of lipopolysaccharide. The above information will be used to better understand the role of individual phospholipids, phospholipid composition and phospholipid content in general cell physiology and specific metabolic processes which will lead to a greater understanding of normal membrane function.